Human induced pluripotent stem cells (hiPSCs) have almost the same characteristics, such as morphology, expandability, surface antigens, gene expression profile, methylation sites, etc., as those of human embryonic stem cells (hESCs). HiPSCs overcome ethical and moral confusions present in hESCs, increase the feasibility of autologous stem cell transplantation therapy, and solve the problem of the limited number of cells.
The culture of human pluripotent stem cells in vitro typically need to rely on mouse embryonic fibroblast (mEFs) or Matrigel. The mEFs culturing system has been gradually substituted by other culturing systems due to its complicated operation processes. Matrigel is a soluble basement membrane extract that obtained from the sarcoma of EHS (Engelbreth-Holm-Swarm) mouse, which can form a layer of thin membrane serving as the extracellular matrix on the surface of a growth substrate and enables human pluripotent stem cells to obtain desired adhesion and self-renewal in in vitro microenvironment established thereby (Xu, C., et al., Feeder-free growth of undifferentiated human embryonic stem cells, Nat Biotechnol, 2001, 19, 10, 971-4). However, since the Matrigel culturing system is heterologous and cannot ensure the consistency of products among different batches and there is an uncertainty of components thereof, it is not suitable for large-scale in vitro expansion of human pluripotent stem cells, which limits its application in clinical therapy. Therefore, a number of scientists have been devoted to the establishment of an in vitro culturing system of human pluripotent stem cells, which may replace Matrigel and has defined components.
First, researchers have sequentially found proteins such as fibronectin, vitronectin, laminin, etc., and certain specific parts of laminin may achieve in vitro self-renewal of human pluripotent stem cells. However, it is difficult for biological products such as proteins, etc., to be widely used in scientific research and clinical practice due to expensive price and batch inconsistency caused by physisorption. Therefore, in recent years, researchers have successfully developed few synthetic surfaces which support in vitro culturing and directional differentiation of human pluripotent stem cells and have significant advantages in terms of cost, batch stability, etc. Villa-Diaz L G et al., have formed a methacrylic acid ester type derivative—PMEDSAH coating—on polystyrene surface as well as a conditioned culture medium or a “StemPro” medium having defined chemical components so as to be capable of achieving the pluripotency maintenance of H9 hESCs (Villa-Diaz L G et al., Synthetic polymer coatings for long-term growth of human embryonic stem cells. Nature biotechnology. 2010; 28:581-3).
By forming a polyacrylic acid layer on the surface of a culture plate and grafting a peptide which promotes adhesion and proliferation of hESCs, Melkoumian Z et al., have experimentally demonstrated that pluripotency maintenance of about 10 passages of hESCs may be achieved on the surface functionalized and modified by the peptide of either Ac-KGGNGEPRGDTYRAY (SEQ ID NO:3) derived from bone sialoprotein (BSP) or Ac-KGGPQVTRGDVFTMP (SEQ ID NO:1) derived from vitronectin (VN) (Melkoumian Z et al., Synthetic peptide-acrylate surfaces for long-term self-renewal and cardiomyocyte differentiation of human embryonic stem cells. Nature biotechnology. 2010; 28:606-10).
However, these synthetic surfaces suffer from the following defects: (1) none has been verified to be capable of supporting somatic cells to be reprogrammed into hiPSCs; (2) most surfaces are not demonstrated to be effectively support a long-term pluripotency maintenance of various human pluripotent stem cells; (3) the production process is complicated; and (4) bio-inert or even bio-toxic components are used. Therefore, it is desirable to establish a simple, stable, and effective in vitro culturing system of human pluripotent stem cells having defined components by using compounds with good biocompatibility.